Globally, 30-40% of new HIV infections occur in adolescents and the HIV epidemic in adolescents is growing. The majority of these infections are acquired via heterosexual intercourse and ~40% of all adolescents living with HIV are boys. It will be impossible to achieve an AIDS-free generation without understanding the adolescent HIV epidemic in boys and developing novel strategies to prevent new infections. Heterosexual transmission of HIV infection occurs across the anogenital mucosa. The presence of HIV target cells in the genital mucosa is a critical determinant of HIV risk. We have shown that penile anaerobes are associated with increased risk of HIV seroconversion, anaerobe abundance correlates with sub-preputial cytokine production, and penile IL-8 is associated with HIV target cell density in the foreskin. Thus, it is biologically plausible that changes in the penile microbiome with resultant inflammatory changes may increase the risk of HIV. There is little information on the pathophysiology of male adolescent HIV infection, especially changes in the mucosa affecting HIV susceptibility associated with maturation and sexual debut. We hypothesize that maturation and sexual debut in boys lead to changes in the penile microbiome that affect the genital immunological milieu and the risk of acquiring HIV and other viral STIs. We propose to evaluate whether the penile microbiome and soluble immunological milieu (secreted cytokines, chemokines and anti-microbial factors) change with reproductive maturation and sexual debut. Using the Rakai Community Cohort Study in Uganda, we will enroll 200 uncircumcised, sexually nave, HIV- negative adolescent boys aged 15-16 years. Urine testosterone and the Tanner scale will be used to assess adolescent maturation. Coronal sulcus penile swabs will be evaluated for the penile microbiome, pro- inflammatory cytokines and chemokines, and specific innate anti-microbial factors (e.g., ?-defensins, ?- defensins, cathelicidins, SLPI and trappin-2/elafin). We will also evaluate the impact of maturation and sexual debut on the tissue immune milieu and HIV susceptibility among the 200 boys in the study who request male circumcision (n~60). Their foreskin tissue and PBMCs will be assessed for inflammatory status, HIV target cell density, microstructural changes and HIV susceptibility using two ex vivo infectivity models. Data will be correlated with the microbiome, inflammation status, HIV/STIs acquisition, sexual debut and maturation stage, and adjusted for HIV risk factors using multivariable regression. This proposal addresses the Request for Applications aims by assessing the genital microbial and immunologic environment that impacts HIV susceptibility in adolescent boys during maturation and sexual debut. This study will contribute to our understanding of host-microbe/immunologic interactions and may lead to novel preventive interventions to reduce HIV risk that could likely be applicable to all men regardless of age, such as targeting high-risk bacterial species with narrow-spectrum antimicrobials or anti-inflammatory agents.